Modified Project Summary/Abstract section Major depressive disorder is a prevalent disease that is debilitating, recurring, progressive, and often inadequately treated with available medications. A better understanding of mechanisms contributing to depression is critically needed to develop new interventions. In this project, we will investigate the influence of microbiota on depressive-like behaviors in mice. Recent evidence has shown that alterations of the gut microbiota influence responses to stress. Microbiota also influences immune responses, in particular certain bacteria regulate the production of T helper (Th) 17 cells. We recently showed that Th17 cells are required for the induction of depressive-like behaviors. Therefore, our overall hypotheses are that stress that induces depression-like behaviors in mice alters signals generated by certain residents of the microflora, that these signals promote the production of pathogenic Th17 cells, which in turn mediate the induction of depression-like behaviors after stress, and that establishment of depression augments changes in the microbiota that facilitates the continued susceptibility to depression, in part through up-regulated production of Th17 cells. In Specific Aim 1, we will test the hypothesis that the microbiota regulates stress-induced pathogenic Th17 cells that promote depressive-like behaviors in mice. We will expand our proof-of-concept data that modulation of the microbiota affects depressive-like behavior and influences Th17 cells in mouse brain during depressive-like states. We identified bacteria that are increased during depressive-like behavior and will confirm their role in controlling depressive-like behavior and Th17 cells production. In Specific Aim 2, we will test the hypothesis that in mice bacterial quorum-sensing molecules are signals that regulate the production of Th17 cells, which promote depression-like behaviors. We generated proof-of-concept data identifying the importance of the quorum sensing molecule AI-2 in predicting depressive-like behavior. We will expand these results by developing therapeutic intervention strategies to modulate bacterial microflora via AI-2 quorum sensing molecules to regulate Th17 cells and depressive-like behavior. Altogether this project will provide evidence that the microbiota regulates depressive-like behavior through the production of Th17 cells.